: Despite great advances in our understanding of gene regulation, the mechanisms by which genes are expressed differentially in neurons, and silenced elsewhere, remain a central issue for neurobiology. Our discovery of the REST repressor complex that regulates the brain Nav1.2 sodium channel, a major determinant of excitability in nerve, has led to more general insights into control of neuronal differentiation. Understanding of the contribution of components of the REST complex, such as CoREST, to neuronal gene expression helps explain how disorders such as epilepsy and Rett Syndrome may give rise to defects in nerve function. Our first specific aim seeks to rigorously test the idea that a REST/CoREST complex initiates a silencing mechanism that propagates along the chromosome over a large distance to encompass genes lacking REST binding sites. If true, this suggests a model whereby higher order chromosome structure is linked to patterns of gene expression. This model has general implications for mechanisms of gene regulation, as well as for potentially providing insights into the ontogeny of the nervous system. A combination of mRNA profiling, bioinformatics, and chromatin biochemistry will be used to test our model. In Aim 2, we seek to elucidate the roles of REST and the co-repressor, CoREST, in regulating gene expression in neuronal progenitor cells (embryonic stem cell and cortical progenitors) and post mitotic neurons. Genes in progenitors cells destined to differentiate are, unlike terminally differentiated cells, repressed rather than silenced by REST. Using RNAi, chromatin immunoprecipitation and microarray analyses, we will elucidate the mechanism for this repression. REST is absent from post mitotic neurons, yet the promoters of REST-regulated genes, such as neuronal calbindin, are methylated; the methylated DNA likely serves as a repressor platform that permits graded expression levels in vivo. We will identify the co-repressor complex that binds to the putative repressor platform in calbindin and determine whether this mechanism of regulation applies to a larger set of neuronal genes.